Acinetobacter baumannii is a major pathogen that causes hospital-acquired infections in China. As an obligate aerobic Gram-negative bacterium, it is widely distributed in healthcare settings and readily colonizes sites such as the skin, conjunctiva, oral cavity, respiratory tract, gastrointestinal tract, and urogenital tract of hospitalized patients (Luo et al., 2025). Infection with A. baumannii has been shown to lead to a range of complications, including ventilator-associated pneumonia, wound and urinary tract infections, and bacteremia. These further exacerbate the physical burden on patients who are hospitalised (Bian et al., 2021).

Carbapenem-resistant A. baumannii has emerged as a primary pathogen of global concern in the domain of hospital infection control (Hujer et al., 2005). Research indicates that approximately 40% of A. baumannii isolates demonstrated carbapenem resistance in 2021 (Jachowicz-Matczak et al., 2025). In the context of intensive care unit (ICU) settings, the implementation of pathogen surveillance has yielded noteworthy insights into the prevalence of carbapenem-resistant strains of A. baumannii. A comprehensive analysis of 148 isolates revealed a staggering 96.7% confirmation rate, with a significant 90% exhibiting extensively drug-resistant (XDR) characteristics (Ababneh et al., 2025). The emergence of multidrug-resistant strains has been shown to have a significant impact on the complexity of clinical treatment and the risk of therapeutic failure (Falagas et al., 2006).

ST2 (international clone IC2) represents a globally disseminated multidrug-resistant lineage and has been consistently reported as a predominant clone in hospital-acquired infections in China. A nationwide genomic analysis covering 27 provinces/municipalities from 1999 to 2022 demonstrated that ST2 accounted for 87.5% of clinical isolates, highlighting its long-term widespread circulation and sustained transmission advantage in the Chinese healthcare setting (Zhang et al., 2025).

Whole-genome sequencing (WGS) has superseded conventional epidemiological investigation methodologies in public health, evolving into a pivotal instrument for disease surveillance and outbreak response (Gorrie et al., 2022). The utilisation of WGS facilitates comprehensive and precise analysis of pathogen genome sequences, expeditious identification of outbreak origins, transmission pathways, and evolutionary trends. This, in turn, provides a robust foundation for the development of effective control strategies. The analysis of WGS data holds significant potential for applications in bacterial classification and the inference of epidemiological features. Specifically, by analysing specific genes within bacterial genomes, it is possible to accurately classify bacterial isolates and further infer their epidemiological characteristics, such as species identification, the detection of resistance genes, and the screening of virulence genes. Research indicates that proactive whole-genome sequencing of healthcare-associated bacterial pathogens significantly reduces morbidity and mortality while generating substantial cost savings for healthcare institutions, thereby offsetting the expenses of implementing genomic surveillance (Fox et al., 2023). The successful dissemination of A. baumannii is closely associated with its high genetic plasticity, which enables long-term persistence in hospital environments and the continuous accumulation of resistance- and virulence-associated genetic traits, thereby facilitating the sustained spread and outbreak formation of high-risk clones (Yehya et al., 2025). As technology advances, the scope and precision of WGS applications continues to expand (Quainoo et al., 2017). The present study employs whole-genome sequencing of A. baumannii to identify the emergence of novel virulence or resistance genes. Phylogenetic analysis is further employed to substantiate the occurrence of outbreaks involving homologous strains.

The objective of this study was to utilise whole-genome sequencing as a means to elucidate the molecular characteristics, antimicrobial resistance profiles, virulence determinants, and phylogenetic relationships of these isolates. The findings of the present study provide critical insights into the long-term dissemination and evolution of A. baumannii in this region, with implications for infection control and antimicrobial stewardship strategies.

A. baumannii is the most prevalent Gram-negative bacillus within the genus Acinetobacter and is an opportunistic pathogen (Humphries et al., 2021). In the most recent 2024 WHO BPPL, carbapenem-resistant A. baumannii continues to be classified as a critical priority pathogen, necessitating the acceleration of efforts to develop new drugs and implement control measures (Jesudason et al., 2024; Jachowicz-Matczak et al., 2025). For patients, treating CRAB infections represents a global challenge (Zhou et al., 2024). In the event of the current limited treatment options proving unsuccessful, CRAB infections have the potential to be catastrophic for patients (Eslami et al., 2025). The present study observed a significantly higher incidence of A. baumannii infection in males than in females, consistent with previous reports (Uslan et al., 2007). Furthermore, a significant increase in infection rates was observed in ICUs in comparison to general wards (Tabah et al., 2023). This observation is reasonable, Patients in the ICU ward present with complex and critical conditions, often exhibiting compromised immunity, a history of multiple antimicrobial agents, and frequent invasive procedures. The treatment and recovery of these patients typically necessitates prolonged hospitalisation. The migration of A. baumannii from colonized sites is facilitated by these factors in combination with invasive interventions, thereby transforming it into a pathogenic bacterium. This phenomenon engenders conditions conducive to the proliferation of A. baumannii infections. Patients undergoing tracheal intubation/tracheotomy are often sedated, which can impair their ability to clear airway secretions and increase the risk of infection due to the downward migration of respiratory colonizing bacteria. Research has demonstrated that A. baumannii colonization rates are 68.1% in the nasal vestibule and 12.8% in the pharynx (Wang et al., 2012). In this study, 56.12% of isolates were obtained from sputum and 15.31% from bronchial lavage fluid, indicating that the respiratory tract is a common site of A. baumannii colonization and infection.

Molecular epidemiology is crucial for understanding the relatedness, evolutionary dynamics, and transmission of different Acinetobacter baumannii lineages, making MLST and whole-genome–based typing indispensable tools. Globally, ST1, ST2, ST25, and ST79 represent the major epidemic lineages of A. baumannii (Hamidian and Nigro, 2019), with ST2 being the dominant clone in China (Li et al., 2019). In the present study, ST2 was likewise identified as the predominant sequence type, consistent with large-scale national and international surveillance data (Duan et al., 2024; Fonseca et al., 2023). Recent phylogenomic analyses have further demonstrated that ST2 is not a sporadically emerging lineage, but rather originated in China in the mid-1970s and subsequently underwent multiple waves of clonal expansion in hospital settings, ultimately becoming a long-standing and persistently circulating dominant lineage (Zhang et al., 2025). In line with this evolutionary and epidemiological context, our analysis further revealed evidence of potential intrahospital transmission of ST2 A. baumannii in our center, characterized by tight genomic clustering and spatiotemporal aggregation of isolates. The high degree of genomic conservation (0–10 SNPs) among ST2 isolates across nearly a decade of sampling is striking. This pattern suggests that rather than undergoing rapid diversifying evolution, the ST2 population in our facility has undergone purifying selection, maintaining a highly optimized genomic architecture. The evolution of this clone appears to be characterized by the acquisition and subsequent fixation of modular resistance determinants (e.g., bla_OXA-23, armA), leading to a stable, dominant ‘MDR backbone’ that is evolutionarily superior to sporadic non-ST2 lineages.

In China, the strong association between ST2 and carbapenem resistance has been systematically documented. A large-scale genomic study of 2,861 A. baumannii isolates collected in China between 1999 and 2022 showed that 96.9% of ST2 strains carried the acquired carbapenemase gene bla_OXA-23 and 98.9% harbored the intrinsic bla_OXA-66, whereas the prevalence of these two genes among non-ST2 isolates was only 34.3 and 13.4%, respectively (Zhang et al., 2025). These national-level data are highly consistent with our findings: in our ST2 isolates, the detection rates of bla_OXA-23 and bla_OXA-66 reached 93.85 and 96.92%, respectively, and the carbapenem resistance rate of ST2 was as high as 95.38%, markedly exceeding that observed in other sequence types. Further genotypic analysis revealed that ST2 strains not only stably carry OXA-type carbapenemases but also systematically accumulate multiple additional resistance determinants, including the aminoglycoside-modifying enzymes aph(3′)-Ia, aph(3′′)-Ib and aph(6)-Id, the sulfonamide and tetracycline resistance genes sul1, sul2 and tet(B), as well as genes mediating high-level aminoglycoside and macrolide resistance such as armA, mphE and msrE. This “modular resistance gene combination” indicates that ST2 has evolved into a highly stable genetic background carrying clustered multidrug-resistance determinants. In contrast, non-ST2 lineages (e.g., ST33 and ST77) display much greater phenotypic and genotypic heterogeneity, with some strains remaining susceptible to multiple antibiotics and exhibiting substantially lower rates of resistance gene carriage. Notably, large-scale genomic studies have shown that the dominance of ST2 is not merely the result of antibiotic selection pressure, but instead reflects a broader evolutionary strategy characterized by extensive recombination and continuous optimization of resistance- and virulence-associated loci, including surface polysaccharides and iron acquisition systems (Liu et al., 2022; Tascini et al., 2025). This evolutionary plasticity allows ST2 to maintain high-level carbapenem resistance while simultaneously enhancing its fitness, persistence, and transmissibility in hospital environments. In this context, the consistently high prevalence of carbapenem resistance (>85%) reported for ST2 worldwide (Duan et al., 2024; Gu et al., 2024) should be interpreted as the outcome of sustained clonal expansion of a biologically optimized, outbreak-prone lineage, rather than independent emergence of resistance in multiple genetic backgrounds.

In addition to its close association with carbapenem resistance, ST2 strains are also recognized for their elevated virulence. Studies have shown that ST2 isolates frequently harbor an extensive arsenal of virulence factors, including genes involved in biofilm formation, iron acquisition systems (such as those encoding siderophores like acinetobactin), and secretion systems, which enhance their ability to survive in hostile environments, evade host immunity, and establish persistent infections (Lurie-Weinberger et al., 2025). This combination of high resistance and enhanced virulence likely contributes to the successful dissemination and poor clinical outcomes associated with ST2 A. baumannii clones. Consistent with this, our study revealed a marked enrichment of iron acquisition genes in ST2 Acinetobacter baumannii. In particular, the complete acinetobactin biosynthesis cluster (basA–basJ) and the transport system (bauB–bauF) were universally conserved, entE was present in all isolates, and the outer membrane receptor bauA was detected in 87.7% of ST2 strains, indicating a highly intact and efficient siderophore-mediated iron uptake system. Because iron is strictly limited in the host, iron acquisition is a key determinant of A. baumannii virulence, and acinetobactin has been shown to be essential for in vivo survival and pathogenicity (Yehya et al., 2025). This optimized iron-scavenging capacity likely promotes bacterial fitness in hospital and host environments, providing a mechanistic basis for the tight SNP clustering, outbreak-prone behavior, and poor clinical outcomes associated with ST2 clones. Importantly, the strong dependence of ST2 on siderophore-mediated iron uptake also highlights these systems as promising therapeutic targets, both through direct inhibition of metallophore pathways and through Trojan-horse antibiotic strategies that exploit siderophore receptors to deliver drugs into multidrug-resistant cells (Ezzeddine and Ghssein, 2023; Tillotson, 2016).

All CRAB strains in this study were multidrug-resistant, with the majority belonging to the highly virulent ST2 clone. In contrast, non-MDR strains exhibited greater genetic diversity (Chen et al., 2024). Pan-genome analysis further highlighted the genomic plasticity of the isolates, with accessory genomes (including shell and cloud genes) accounting for 76.4% of the total gene pool—significantly larger than the core genome (23.6%). This suggests a high degree of genetic mobility and adaptive potential, often associated with the acquisition of antibiotic resistance genes, virulence determinants, and mobile genetic elements. Such genomic flexibility may underpin the successful expansion and persistence of ST2 strains in hospital settings in Guangzhou (Galac et al., 2020; Milani et al., 2021). The open pan-genome structure of A. baumannii, continuously integrating new accessory genes, likely facilitates its adaptation to diverse clinical environments (Ali et al., 2022). The convergence of genomic plasticity and the multidrug-resistant phenotype observed in this study emphasizes the need for strengthened surveillance and control measures against high-risk clones in healthcare settings.

This was a single-center retrospective study, which may limit the generalizability of the observed epidemiological and genomic patterns of A. baumannii. In addition, because no ST2 isolates were available from 2013, the earliest phase of ST2 introduction into our hospital could not be directly evaluated. Although strong genomic clustering and ward-level aggregation of ST2 isolates were identified, the absence of environmental and healthcare worker sampling prevented precise reconstruction of transmission routes. Moreover, the use of short-read sequencing and database-based annotation may have limited the detection of novel resistance determinants, plasmid-mediated transmission, and structural genomic variations. Future studies should therefore integrate multicenter prospective surveillance, long-read sequencing, environmental sampling, and real-time clinical metadata to better resolve ST2 transmission networks and support early detection and targeted infection control of high-risk clones.

In conclusion, this retrospective genomic epidemiological study provides a comprehensive characterisation of Acinetobacter baumannii clinical isolates from a tertiary hospital in Guangzhou over a nine-year period. The findings of this study serve to emphasise the ongoing predominance of the ST2 clone, which demonstrated elevated levels of carbapenem resistance and an extensive array of antimicrobial resistance genes, including bla_OXA-23 and bla_OXA-66. These factors are likely to have contributed to its effective nosocomial propagation. The high prevalence of resistance to multiple antibiotic classes is indicative of the critical challenge posed by CRAB in clinical settings. Furthermore, the pan-genome analysis revealed a highly plastic accessory genome, facilitating rapid adaptation and resistance acquisition. The prevalence of virulence factors associated with iron acquisition and biofilm formation further elucidates the pathogenic potential of these isolates. These results emphasise the urgent need for strengthened infection control measures, antimicrobial stewardship programs, and continuous genomic surveillance to monitor the evolution and spread of high-risk clones. It is recommended that future studies concentrate on the functional validation of resistance and virulence mechanisms, as well as the role of mobile genetic elements in the horizontal transfer of resistance genes. The objective of this research is to inform targeted therapeutic and preventive strategies.